Plateforme de Biophysique Moléculaire, UMR 3528 CNRS, Institut Pasteur, 25 Rue du docteur Roux, Cedex 15, 75724, Paris, France.
Eur Biophys J. 2021 May;50(3-4):587-595. doi: 10.1007/s00249-020-01492-3. Epub 2021 Jan 24.
Intrinsic viscosity is a key hydrodynamic parameter to understand molecular structure and hydration, as well as intramolecular interactions. Commercially available instruments measure intrinsic viscosity by recording the macromolecular mobility in a capillary. These instruments monitor Taylor dispersion using an absorbance or fluorescence detector. By design, these instruments behave like U-tube viscometers. To our knowledge, there are no studies to date showing that the Viscosizer TD instrument (Malvern-Panalytical) is able to measure the intrinsic viscosity of macromolecules. In this study, we then performed our assays on the Poly(ethylene oxide) polymer (PEO), used classically as a standard for viscometry measurements and on three model proteins: the bovine serum albumin (BSA), the bevacizumab monoclonal antibody, and the RTX Repeat Domain (RD) of the adenylate cyclase toxin of Bordetella pertussis (CyaA). The presence of P20 in the samples is critical to get reliable results. The data obtained with our in-house protocol show a strong correlation with intrinsic viscosity values obtained using conventional techniques. However, with respect to them, our measurements could be performed at relatively low concentrations, between 2 and 5 mg/ml, using only 7 µL per injection. Altogether, our results show that the Viscosizer TD instrument is able to measure intrinsic viscosities in a straightforward manner. This simple and innovative approach should give a new boost to intrinsic viscosity measurements and should reignite the interest of biophysicists, immunologists, structural biologists and other researchers for this key physicochemical parameter.
特性黏度是理解分子结构和水合作用以及分子内相互作用的关键流体力学参数。市售仪器通过记录毛细管中大分子的迁移率来测量特性黏度。这些仪器使用吸光度或荧光检测器监测泰勒分散。根据设计，这些仪器的行为类似于 U 型管粘度计。据我们所知，目前尚无研究表明 Viscosizer TD 仪器（马尔文帕纳分析公司）能够测量大分子的特性黏度。在这项研究中，我们随后对聚（环氧乙烷）聚合物（PEO）进行了测定，PEO 通常用作粘度测量的标准，还对三种模型蛋白进行了测定：牛血清白蛋白（BSA）、贝伐单抗单克隆抗体和百日咳博德特氏菌腺苷酸环化酶毒素的 RTX 重复结构域（CyaA）。样品中 P20 的存在对于获得可靠的结果至关重要。我们的内部方案获得的数据与使用传统技术获得的特性黏度值具有很强的相关性。然而，与它们相比，我们的测量可以在相对较低的浓度（2 至 5mg/ml）下进行，每次注射仅需 7µL。总的来说，我们的结果表明 Viscosizer TD 仪器能够以简单直接的方式测量特性黏度。这种简单而创新的方法应该会对特性黏度测量产生新的推动作用，并重新激发生物物理学家、免疫学家、结构生物学家和其他研究人员对这一关键物理化学参数的兴趣。
